Hydrophobic interactions of the structural protein GRP1.8 in the cell wall of protoxylem elements. (25/376)

The glycine-rich structural protein GRP1.8 of French bean (Phaseolus vulgaris) is specifically localized in the modified primary cell walls of protoxylem elements. Continuous deposition of GRP1.8 into the cell walls during elongation growth of the plant suggests that GRP1.8 is part of a repair mechanism to strengthen the protoxylem. In this work, a reporter-protein system was developed to study the interaction of GRP1.8 with the extracellular matrix. Fusion proteins of a highly soluble chitinase with different domains of GRP1.8 were expressed in the vascular tissue of tobacco (Nicotiana tabacum), and the chemical nature of the interaction of these fusion proteins in the cell wall compartment was analyzed. In contrast with chitinase that required only low-salt conditions for complete extraction, the different chitinase/GRP1.8 fusion proteins were completely extracted only by a nonionic or ionic detergent, indicating hydrophobic interactions of GRP1.8. The same interactions were found with the endogenous GRP1.8 in bean hypocotyls. In addition, in vitro experiments indicate that oxidative cross-linking of tyrosines might account for the insolubilization of GRP1.8 observed in later stages of protoxylem development. Our data suggest that GRP1.8 forms a hydrophobic protein-layer in the cell wall of protoxylem vessels.  (+info)

Sex determination in the monoecious species cucumber is confined to specific floral whorls. (26/376)

In unisexual flowers, sex is determined by the selective repression of growth or the abortion of either male or female reproductive organs. The mechanism by which this process is controlled in plants is still poorly understood. Because it is known that the identity of reproductive organs in plants is controlled by homeotic genes belonging to the MADS box gene family, we analyzed floral homeotic mutants from cucumber, a species that bears both male and female flowers on the same individual. To study the characteristics of sex determination in more detail, we produced mutants similar to class A and C homeotic mutants from well-characterized hermaphrodite species such as Arabidopsis by ectopically expressing and suppressing the cucumber gene CUCUMBER MADS1 (CUM1). The cucumber mutant green petals (gp) corresponds to the previously characterized B mutants from several species and appeared to be caused by a deletion of 15 amino acid residues in the coding region of the class B MADS box gene CUM26. These homeotic mutants reveal two important concepts that govern sex determination in cucumber. First, the arrest of either male or female organ development is dependent on their positions in the flower and is not associated with their sexual identity. Second, the data presented here strongly suggest that the class C homeotic function is required for the position-dependent arrest of reproductive organs.  (+info)

Importance of the N-terminal sequence of the extrinsic 23 kDa polypeptide in Photosystem II in ion retention in oxygen evolution. (27/376)

The function of the extrinsic 23 kDa polypeptide (OEC23) in Photosystem II (PS II) is to retain Ca(2+) and Cl(-) during the S-state turnover of the Mn cluster in photosynthetic oxygen evolution. Recombinant OEC23s from several plant species were produced in Escherichia coli to characterize the molecular mechanism of the OEC23 function then used in reconstitution experiments. One tobacco isoform, OEC23 (2AF), had much less oxygen-evolving activity than the spinach and cucumber OEC23s when PS II activities were reconstituted in salt-washed spinach PS II particles. The fact that the OEC23s had similar binding affinities for PS II particles suggests different ion-retention capacities for the individual OEC23s: The chimeric OEC23s produced between spinach and 2AF and those produced between cucumber and 2AF show that 58 N-terminal amino acid residues are important for PS II activity. Further dissection of the sequence and site-directed mutagenesis indicated the importance of 20 N-terminal amino acid residues for activity, in particular the asparagine at the 15th position. In spinach the N15D mutation decreased PS II activity, whereas in 2AF the D15N mutation increased it. This shows the importance of the N-terminal sequence of OEC23 in ion retention during the water-splitting process.  (+info)

Generation of a murine single chain Fv (scFv) antibody specific for cucumber mosaic virus (CMV) using a phage display library. (28/376)

With the long-term goal of generating CMV-resistant transgenic plants using antibody genes, a single-chain variable fragment (scFv) antibody that binds to the cucumber mosaic virus was isolated from a scFv phage display library by four rounds of affinity selection with CMV-Mf as an antigen. The scFv has the identical binding specificity to CMV as a monoclonal antibody that is generated by the hybridoma fusion technique, and recognized purified preparations of CMV isolates belonging to either subgroup I or II in immunoblotting. The nucleotide sequences of the recombinant antibody showed that a heavy chain variable region (V(H)) gene belonged to the VH3 subgroup and the kappa light chain variable region (V kappa) came from the Vkappa4 subgroup. Our results demonstrate that the scFv phage display library, an alternative approach to the traditional hybridoma fusion technique, has a potential applicability in the study of plant virus and plant pathology.  (+info)

Impact of biocontrol Pseudomonas fluorescens CHA0 and a genetically modified derivative on the diversity of culturable fungi in the cucumber rhizosphere. (29/376)

Little is known about the effects of Pseudomonas biocontrol inoculants on nontarget rhizosphere fungi. This issue was addressed using the biocontrol agent Pseudomonas fluorescens CHA0-Rif, which produces the antimicrobial polyketides 2,4-diacetylphloroglucinol (Phl) and pyoluteorin (Plt) and protects cucumber from several fungal pathogens, including Pythium spp., as well as the genetically modified derivative CHA0-Rif(pME3424). Strain CHA0-Rif(pME3424) overproduces Phl and Plt and displays improved biocontrol efficacy compared with CHA0-Rif. Cucumber was grown repeatedly in the same soil, which was left uninoculated, was inoculated with CHA0-Rif or CHA0-Rif(pME3424), or was treated with the fungicide metalaxyl (Ridomil). Treatments were applied to soil at the start of each 32-day-long cucumber growth cycle, and their effects on the diversity of the rhizosphere populations of culturable fungi were assessed at the end of the first and fifth cycles. Over 11,000 colonies were studied and assigned to 105 fungal species (plus several sterile morphotypes). The most frequently isolated fungal species (mainly belonging to the genera Paecilomyces, Phialocephala, Fusarium, Gliocladium, Penicillium, Mortierella, Verticillium, Trichoderma, Staphylotrichum, Coniothyrium, Cylindrocarpon, Myrothecium, and Monocillium) were common in the four treatments, and no fungal species was totally suppressed or found exclusively following one particular treatment. However, in each of the two growth cycles studied, significant differences were found between treatments (e.g., between the control and the other treatments and/or between the two inoculation treatments) using discriminant analysis. Despite these differences in the composition and/or relative abundance of species in the fungal community, treatments had no effect on species diversity indices, and species abundance distributions fit the truncated lognormal function in most cases. In addition, the impact of treatments at the 32-day mark of either growth cycle was smaller than the effect of growing cucumber repeatedly in the same soil.  (+info)

Growth interactions during bacterial colonization of seedling rootlets. (30/376)

Rootlet elongation and bacterial growth on rootlets were determined after inoculation of cucumber and spinach seedlings with Pseudomonas strains differing in production of siderophores and HCN. Siderophore producers grew more profusely than nonproducers on both species and promoted rootlet elongation on cucumber. Coinoculation of siderophore producers and nonproducers resulted in restricted growth of the latter. The total populations of nonproducers of HCN in the presence of HCN producers were not decreased, but the tenacity of their association with the rootlet surface was altered.  (+info)

Expression pattern of the CsPK3 auxin-responsive protein kinase gene. (31/376)

We have previously cloned a cDNA of a putative serine/threonine protein kinase gene named CsPK3 from cucumber, the mRNA level of which was up-regulated by auxin and down-regulated by light irradiation. To examine the CsPK3 gene expression in detail, we cloned a genomic DNA of CsPK3 gene and made transgenic tobacco (Nicotiana tabacum L. cv. Petit Havana SR1) plants containing the fused CsPK3 promoter-beta-glucuronidase gene. The beta-glucuronidase expression was detected in the shoot apex, vascular tissues, and the outermost layer of cortex. The histological distribution of CsPK3 mRNA in cucumber seedlings was supported by in situ hybridization, where the positive signals were observed in similar tissues as those observed by beta-glucuronidase staining. The responsiveness of the CsPK3 gene to auxin and light was also confirmed for beta-glucuronidase activity. The pattern of beta-glucuronidase staining changed during the development of the tobacco seedlings. The results of our experiment showed that CsPK3 was expressed in a wide variety of tissues and cells in which the developmental and growth controls by auxin are suggested.  (+info)

Identification of specific cucumber necrosis virus coat protein amino acids affecting fungus transmission and zoospore attachment. (32/376)

Cucumber necrosis virus (CNV) is naturally transmitted in the soil by zoospores of the fungal vector Olpidium bornovanus. Successful transmission requires that virus particles attach to the surface of zoospores prior to zoospore encystment on host roots. Mechanically passaged CNV was screened for mutants deficient in fungus transmission. We found six such mutants, exhibiting transmission efficiencies ranging from approximately 14 to 76% of that of wild-type (WT) CNV. Results of in vitro virus-zoospore binding assays show that each mutant binds to zoospores less efficiently than WT CNV (21 to 68%), suggesting that defects in transmission for these mutants are at least partially due to inefficient zoospore binding. Analysis of the structure of the CNV coat protein subunit and trimer indicates that affected amino acids in all of the mutants are located in the shell or protruding domain and that five of six of them are potentially exposed on the surface of the virus particle. In addition, several of the mutated sites, along with a previously identified site in a region of subunit-subunit interaction in the coat protein shell domain (M. A. Robbins, R. D. Reade, and D. M. Rochon, Virology 234:138-146, 1997), are located on the particle quasi-threefold axis, suggesting that this region of the capsid may be important in recognition of a putative zoospore receptor. The individual sites may directly affect attachment to a receptor or could indirectly affect attachment via changes in virion conformation.  (+info)